Effect of microRNA-543 by regulating RNA-binding motif proteins 47 expression on proliferation and metastasis of gastric cancer cells
Objective To explore the effect of microRNA-543(miR-543)via inhibiting RNA-binding motif proteins 47(RBM47)on proliferation and metastasis in gastric cancer(GC)and its un-derlying mechanism.Methods A total of 40 specimens of gastric cancer tissues and normal tissues were collected,and the level of miR-543 was tested by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR).Normal gastric epithelial cell line GES-1,and gastric cancer cell lines(MKN45,NCIN87 and AGS)were cultured,and the expression of miR-543 was detected by RT-qPCR.The correlation between the clinicopathological characteristics and miR-543 was analyzed.Bioinformatics a-nalysis was used to detect the target genes of miR-543 in GC.KEGG and GO analyses were carried out to explore the regulatory functions of miR-543.The target of miR-543 found by bioinformatics analysis was verified by luciferase reporter assay.Moreover,after miR-543 inhibitor was transfected into AGS cells,the proliferation,migration,and invasion of GC cells were detected by cell counting kit-8(CCK-8)assay,wound healing assay,and Transwell assay.The expression of Cyclin D1,matrix metalloprotein-9(MMP-9),RBM47,and tissue inhibitor of metal protease 1(TIMP1)was tested in AGS before and after the transfection by RT-qPCR and Western blotting.SPSS 25.0 was used to analyze the data.ResultsThe level of miR-543 was higher in GC tissues(2.476±0.812)than that in normal tissues(1.165±0.295,t=9.880,P＜0.01),which was further confirmed by RT-qPCR results in GC cells,indicating miR-543 had higher expression in GC cells than in normal gastric epithelial cells(1.130±0.141,t=9.842,P＜0.05;t=13.17,P＜0.01;t=35.07,P＜0.01),and AGS cell line had the highest level of miR-543(5.120±0.545).Meanwhile,miR-543 was correlated with lymph node metastasis(positive cases:27).Functional analysis proved that miR-543 participated in various processes including intracellular protein transport,protein ubiquitination,and apoptotic process,and miR-543 targeted at RBM47.RBM47 was downregulated in GC(0.663±0.251)as compared with normal tissues(1.604±0.613,P＜0.01)and correlation analysis showed that miR-543 was negatively related to RBM47(r=-0.724),which was fur-ther verified by luciferase reporter assay.After silencing miR-543,the proliferation(P＜0.01),migration and invasion of GC cells were inhibited significantly,and the expression of Cyclin D1 and MMP-9 was de-creased(P＜0.01)while the expression of RBM47 and TIMP1 was increased(P＜0.01)in miR-543 in-hibitor group.Conclusion MiR-543 contributes to the proliferation and metastasis of GC cells by down-regulating RBM47 level.
NETL
NSTL
万方数据
